{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,28]],"date-time":"2026-02-28T04:29:53Z","timestamp":1772252993250,"version":"3.50.1"},"reference-count":49,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2020,3,18]],"date-time":"2020-03-18T00:00:00Z","timestamp":1584489600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100000268","name":"Biotechnology and Biological Sciences Research Council","doi-asserted-by":"publisher","award":["BB\/E\/I\/00007035"],"award-info":[{"award-number":["BB\/E\/I\/00007035"]}],"id":[{"id":"10.13039\/501100000268","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000268","name":"Biotechnology and Biological Sciences Research Council","doi-asserted-by":"publisher","award":["BB\/E\/I\/00007036"],"award-info":[{"award-number":["BB\/E\/I\/00007036"]}],"id":[{"id":"10.13039\/501100000268","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000268","name":"Biotechnology and Biological Sciences Research Council","doi-asserted-by":"publisher","award":["BBS\/E\/I\/00007039"],"award-info":[{"award-number":["BBS\/E\/I\/00007039"]}],"id":[{"id":"10.13039\/501100000268","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000268","name":"Biotechnology and Biological Sciences Research Council","doi-asserted-by":"publisher","award":["BBS\/E\/I\/00001942"],"award-info":[{"award-number":["BBS\/E\/I\/00001942"]}],"id":[{"id":"10.13039\/501100000268","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000268","name":"Biotechnology and Biological Sciences Research Council","doi-asserted-by":"publisher","award":["BB\/K011057\/1"],"award-info":[{"award-number":["BB\/K011057\/1"]}],"id":[{"id":"10.13039\/501100000268","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000199","name":"U.S. Department of Agriculture","doi-asserted-by":"publisher","award":["6040-32000-074-00D"],"award-info":[{"award-number":["6040-32000-074-00D"]}],"id":[{"id":"10.13039\/100000199","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>Marek\u2019s disease is a major scourge challenging poultry health worldwide. It is caused by the highly contagious Marek\u2019s disease virus (MDV), an alphaherpesvirus. Here, we showed that, similar to other members of its Herpesviridae family, MDV also presents a complex landscape of splicing events, most of which are uncharacterised and\/or not annotated. Quite strikingly, and although the biological relevance of this fact is unknown, we found that a number of viral splicing isoforms are strain-specific, despite the close sequence similarity of the strains considered: very virulent RB-1B and vaccine CVI-988. We validated our findings by devising an assay that discriminated infections caused by the two strains in chicken embryonic fibroblasts on the basis of the presence of some RNA species. To our knowledge, this study is the first to accomplish such a result, emphasizing how relevant a comprehensive picture of the viral transcriptome is to fully understand viral pathogenesis.<\/jats:p>","DOI":"10.3390\/v12030329","type":"journal-article","created":{"date-parts":[[2020,3,19]],"date-time":"2020-03-19T03:54:14Z","timestamp":1584590054000},"page":"329","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Pervasive Differential Splicing in Marek\u2019s Disease Virus Can Discriminate CVI-988 Vaccine Strain from RB-1B Very Virulent Strain in Chicken Embryonic Fibroblasts"],"prefix":"10.3390","volume":"12","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1892-1361","authenticated-orcid":false,"given":"Yashar","family":"Sadigh","sequence":"first","affiliation":[{"name":"Avian Viral Oncogenesis, The Pirbright Institute, Ash Road, Woking GU24 0NF, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Abdessamad","family":"Tahiri-Alaoui","sequence":"additional","affiliation":[{"name":"Clinical BioManufacturing Facility, The Jenner Institute, University of Oxford, Old Road, Headington, Oxford OX3 7JT, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Stephen","family":"Spatz","sequence":"additional","affiliation":[{"name":"US National Poultry Research Center, 934 College Station Road, Athens, GA 30605, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8255-516X","authenticated-orcid":false,"given":"Venugopal","family":"Nair","sequence":"additional","affiliation":[{"name":"Avian Viral Oncogenesis, The Pirbright Institute, Ash Road, Woking GU24 0NF, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5599-3933","authenticated-orcid":false,"given":"Paolo","family":"Ribeca","sequence":"additional","affiliation":[{"name":"Integrative Biology and Bioinformatics, The Pirbright Institute, Ash Road, Woking GU24 0NF, UK"},{"name":"Biomathematics and Statistics Scotland (BioSS), James Clerk Maxwell Building, Peter Guthrie Tait Road, The King\u2019s Buildings, Edinburgh EH9 3FD, UK"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2020,3,18]]},"reference":[{"key":"ref_1","unstructured":"Swayne, D., Glisson, J., McDougald, L., Nolan, L., Suarez, D., and Nair, V. (2013). Neoplastic disease: Marek\u2019s disease. Diseases of Poultry, Wyley-Blackwell. [13th ed.]."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"78","DOI":"10.1016\/j.vetimm.2006.03.014","article-title":"Vaccinal control of Marek\u2019s disease: Current challenges, and future strategies to maximize protection","volume":"112","author":"Baigent","year":"2006","journal-title":"Vet. Immunol. Immunopathol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"113","DOI":"10.1016\/j.vetmic.2016.11.024","article-title":"Marek\u2019s disease vaccines: Current status, and strategies for improvement and development of vector vaccines","volume":"206","author":"Reddy","year":"2017","journal-title":"Vet. Microbiol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"126","DOI":"10.2307\/1588906","article-title":"Control of Marek\u2019s disease in the Netherlands. II. Field trials on vaccination with an avirulent strain (CVI 988) of Marek\u2019s disease virus","volume":"16","author":"Rispens","year":"1972","journal-title":"Avian Dis."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"108","DOI":"10.2307\/1588905","article-title":"Control of Marek\u2019s disease in the Netherlands. I. Isolation of an avirulent Marek\u2019s disease virus (strain CVI 988) and its use in laboratory vaccination trials","volume":"16","author":"Rispens","year":"1972","journal-title":"Avian Dis."},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Read, A.F., Baigent, S.J., Powers, C., Kgosana, L.B., Blackwell, L., Smith, L.P., Kennedy, D.A., Walkden-Brown, S.W., and Nair, V.K. (2015). Imperfect Vaccination Can Enhance the Transmission of Highly Virulent Pathogens. PLoS Biol., 13.","DOI":"10.1371\/journal.pbio.1002198"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"149","DOI":"10.1016\/j.tvjl.2004.10.005","article-title":"Marek\u2019s disease: The continuing struggle between pathogen and host","volume":"170","author":"Witter","year":"2005","journal-title":"Vet. J."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"S46","DOI":"10.1080\/03079459808419292","article-title":"The changing landscape of Marek\u2019s disease","volume":"27","author":"Witter","year":"1998","journal-title":"Avian Pathol"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"153","DOI":"10.1637\/11525-110216-Reg.1","article-title":"Industry-Wide Surveillance of Marek\u2019s Disease Virus on Commercial Poultry Farms","volume":"61","author":"Kennedy","year":"2017","journal-title":"Avian Dis."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"167","DOI":"10.1016\/j.rvsc.2010.08.002","article-title":"Differential quantification of cloned CVI988 vaccine strain and virulent RB-1B strain of Marek\u2019s disease viruses in chicken tissues, using real-time PCR","volume":"91","author":"Baigent","year":"2011","journal-title":"Res. Vet. Sci."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"23","DOI":"10.1016\/j.jviromet.2016.03.002","article-title":"Real-time PCR for differential quantification of CVI988 vaccine virus and virulent strains of Marek\u2019s disease virus","volume":"233","author":"Baigent","year":"2016","journal-title":"J. Virol. Methods"},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Tombacz, D., Balazs, Z., Csabai, Z., Moldovan, N., Szucs, A., Sharon, D., Snyder, M., and Boldogkoi, Z. (2017). Characterization of the Dynamic Transcriptome of a Herpesvirus with Long-read Single Molecule Real-Time Sequencing. Sci. Rep., 7.","DOI":"10.1038\/srep43751"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1079","DOI":"10.3389\/fmicb.2017.01079","article-title":"Long-Read Isoform Sequencing Reveals a Hidden Complexity of the Transcriptional Landscape of Herpes Simplex Virus Type 1","volume":"8","author":"Tombacz","year":"2017","journal-title":"Front. Microbiol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1088","DOI":"10.1126\/science.1227919","article-title":"Decoding human cytomegalovirus","volume":"338","author":"Weisburd","year":"2012","journal-title":"Science"},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Arias, C., Weisburd, B., Stern-Ginossar, N., Mercier, A., Madrid, A.S., Bellare, P., Holdorf, M., Weissman, J.S., and Ganem, D. (2014). KSHV 2.0: A comprehensive annotation of the Kaposi\u2019s sarcoma-associated herpesvirus genome using next-generation sequencing reveals novel genomic and functional features. PLoS Pathog., 10.","DOI":"10.1371\/journal.ppat.1003847"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"437","DOI":"10.1038\/nrm.2017.27","article-title":"Alternative splicing as a regulator of development and tissue identity","volume":"18","author":"Baralle","year":"2017","journal-title":"Nat. Rev. Mol. Cell Biol."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Huertas, C.S., Dominguez-Zotes, S., and Lechuga, L.M. (2017). Analysis of alternative splicing events for cancer diagnosis using a multiplexing nanophotonic biosensor. Sci. Rep., 7.","DOI":"10.1038\/srep41368"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"6623","DOI":"10.1007\/s13277-015-3865-5","article-title":"Survivin splice variants and their diagnostic significance","volume":"36","author":"Sah","year":"2015","journal-title":"Tumour Biol."},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Leman, R., Gaildrat, P., Le Gac, G., Ka, C., Fichou, Y., Audrezet, M.P., Caux-Moncoutier, V., Caputo, S.M., Boutry-Kryza, N., and Leone, M. (2019). Novel diagnostic tool for prediction of variant spliceogenicity derived from a set of 395 combined in silico\/in vitro studies: An international collaborative effort. Nucleic Acids Res.","DOI":"10.1093\/nar\/gkz1212"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1185","DOI":"10.1038\/nmeth.2221","article-title":"The GEM mapper: Fast, accurate and versatile alignment by filtration","volume":"9","author":"Sammeth","year":"2012","journal-title":"Nature Methods"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"506","DOI":"10.1038\/nature12531","article-title":"Transcriptome and genome sequencing uncovers functional variation in humans","volume":"501","author":"Lappalainen","year":"2013","journal-title":"Nature"},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Derrien, T., Estell\u00e9, J., Marco Sola, S., Knowles, D.G., Raineri, E., Guig\u00f3, R., and Ribeca, P. (2012). Fast computation and applications of genome mappability. PLoS One, 7.","DOI":"10.1371\/journal.pone.0030377"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"4288","DOI":"10.1093\/nar\/gks042","article-title":"Differential expression analysis of multifactor RNA-Seq experiments with respect to biological variation","volume":"40","author":"McCarthy","year":"2012","journal-title":"Nucleic Acids Res."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"1630","DOI":"10.1101\/gr.094607.109","article-title":"JBrowse: A next-generation genome browser","volume":"19","author":"Skinner","year":"2009","journal-title":"Genome Res."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"53","DOI":"10.1016\/j.jviromet.2004.08.019","article-title":"Absolute quantitation of Marek\u2019s disease virus genome copy number in chicken feather and lymphocyte samples using real-time PCR","volume":"123","author":"Baigent","year":"2005","journal-title":"J. Virol Methods"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"e45","DOI":"10.1093\/nar\/29.9.e45","article-title":"A new mathematical model for relative quantification in real-time RT-PCR","volume":"29","author":"Pfaffl","year":"2001","journal-title":"Nucleic Acids Res."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"2708","DOI":"10.3389\/fmicb.2017.02708","article-title":"Multi-Platform Sequencing Approach Reveals a Novel Transcriptome Profile in Pseudorabies Virus","volume":"8","author":"Moldovan","year":"2017","journal-title":"Front. Microbiol."},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Tombacz, D., Csabai, Z., Olah, P., Balazs, Z., Liko, I., Zsigmond, L., Sharon, D., Snyder, M., and Boldogkoi, Z. (2016). Full-Length Isoform Sequencing Reveals Novel Transcripts and Substantial Transcriptional Overlaps in a Herpesvirus. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0162868"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1604","DOI":"10.1128\/JVI.02989-13","article-title":"Global bidirectional transcription of the Epstein-Barr virus genome during reactivation","volume":"88","author":"Cao","year":"2014","journal-title":"J. Virol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"77","DOI":"10.1006\/viro.1996.0629","article-title":"Characterization of the protein product encoded by a splicing variant of the Marek\u2019s disease virus Eco-Q gene (Meq)","volume":"226","author":"Peng","year":"1996","journal-title":"Virology"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"9","DOI":"10.1007\/s11262-006-0004-9","article-title":"Multiple alternative splicing to exons II and III of viral interleukin-8 (vIL-8) in the Marek\u2019s disease virus genome: The importance of vIL-8 exon I","volume":"34","author":"Jarosinski","year":"2007","journal-title":"Virus Genes"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1713","DOI":"10.1099\/0022-1317-75-7-1713","article-title":"Identification of novel transcripts complementary to the Marek\u2019s disease virus homologue of the ICP4 gene of herpes simplex virus","volume":"75 (Pt. 7)","author":"Li","year":"1994","journal-title":"J. Gen. Virol."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"7896","DOI":"10.1128\/JVI.00908-12","article-title":"Marek\u2019s disease virus expresses multiple UL44 (gC) variants through mRNA splicing that are all required for efficient horizontal transmission","volume":"86","author":"Jarosinski","year":"2012","journal-title":"J. Virol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"1348","DOI":"10.1128\/JVI.02687-14","article-title":"The ORF012 gene of Marek\u2019s disease virus type 1 produces a spliced transcript and encodes a novel nuclear phosphoprotein essential for virus growth","volume":"89","author":"Schippers","year":"2015","journal-title":"J. Virol."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"88","DOI":"10.1016\/j.virol.2014.11.008","article-title":"An MHC class I immune evasion gene of Mareks disease virus","volume":"475","author":"Hearn","year":"2015","journal-title":"Virology"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"154","DOI":"10.1016\/j.vetmic.2006.06.019","article-title":"Expression of Marek\u2019s disease virus phosphorylated polypeptide pp38 produces splice variants and enhances metabolic activity","volume":"117","author":"Li","year":"2006","journal-title":"Vet. Microbiol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"7980","DOI":"10.1128\/JVI.74.17.7980-7988.2000","article-title":"The genome of a very virulent Marek\u2019s disease virus","volume":"74","author":"Tulman","year":"2000","journal-title":"J. Virol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"6091","DOI":"10.1073\/pnas.97.11.6091","article-title":"The complete unique long sequence and the overall genomic organization of the GA strain of Marek\u2019s disease virus","volume":"97","author":"Lee","year":"2000","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1080","DOI":"10.1099\/vir.0.82600-0","article-title":"Comparative full-length sequence analysis of oncogenic and vaccine (Rispens) strains of Marek\u2019s disease virus","volume":"88","author":"Spatz","year":"2007","journal-title":"J. Gen. Virol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"3593","DOI":"10.1128\/jvi.68.6.3593-3603.1994","article-title":"Identification of an immediate-early gene in the Marek\u2019s disease virus long internal repeat region which encodes a unique 14-kilodalton polypeptide","volume":"68","author":"Hong","year":"1994","journal-title":"J. Virol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"5159","DOI":"10.1128\/JVI.75.11.5159-5173.2001","article-title":"Marek\u2019s disease virus (MDV) encodes an interleukin-8 homolog (vIL-8): Characterization of the vIL-8 protein and a vIL-8 deletion mutant MDV","volume":"75","author":"Parcells","year":"2001","journal-title":"J. Virol."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"8712","DOI":"10.1128\/JVI.77.16.8712-8718.2003","article-title":"Replication-competent bacterial artificial chromosomes of Marek\u2019s disease virus: Novel tools for generation of molecularly defined herpesvirus vaccines","volume":"77","author":"Petherbridge","year":"2003","journal-title":"J. Virol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"13376","DOI":"10.1128\/JVI.78.23.13376-13380.2004","article-title":"Oncogenicity of virulent Marek\u2019s disease virus cloned as bacterial artificial chromosomes","volume":"78","author":"Petherbridge","year":"2004","journal-title":"J. Virol."},{"key":"ref_44","unstructured":"Davison, T.F., Nair, V., and Institute for Animal Health (Great Britain) (2004). Marek\u2019s Disease: An Evolving Problem, Elsevier."},{"key":"ref_45","doi-asserted-by":"crossref","unstructured":"Tirosh, O., Cohen, Y., Shitrit, A., Shani, O., Le-Trilling, V.T., Trilling, M., Friedlander, G., Tanenbaum, M., and Stern-Ginossar, N. (2015). The Transcription and Translation Landscapes during Human Cytomegalovirus Infection Reveal Novel Host-Pathogen Interactions. PLoS Pathog., 11.","DOI":"10.1371\/journal.ppat.1005288"},{"key":"ref_46","doi-asserted-by":"crossref","unstructured":"Armero, V.E.S., Tremblay, M.P., Allaire, A., Boudreault, S., Martenon-Brodeur, C., Duval, C., Durand, M., Lapointe, E., Thibault, P., and Tremblay-Letourneau, M. (2017). Transcriptome-wide analysis of alternative RNA splicing events in Epstein-Barr virus-associated gastric carcinomas. PLoS ONE, 12.","DOI":"10.1371\/journal.pone.0176880"},{"key":"ref_47","doi-asserted-by":"crossref","unstructured":"Bertzbach, L.D., Pfaff, F., Pauker, V.I., Kheimar, A.M., Hoper, D., Hartle, S., Karger, A., and Kaufer, B.B. (2019). The Transcriptional Landscape of Marek\u2019s Disease Virus in Primary Chicken B Cells Reveals Novel Splice Variants and Genes. Viruses, 11.","DOI":"10.3390\/v11030264"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"593","DOI":"10.1080\/03079458208436134","article-title":"Characterisation of two highly oncogenic strains of Marek\u2019s disease virus","volume":"11","author":"Schat","year":"1982","journal-title":"Avian Pathol."},{"key":"ref_49","doi-asserted-by":"crossref","unstructured":"Conradie, A.M., Bertzbach, L.D., Bhandari, N., Parcells, M., and Kaufer, B.B. (2019). A Common Live-Attenuated Avian Herpesvirus Vaccine Expresses a Very Potent Oncogene. mSphere, 4.","DOI":"10.1128\/mSphere.00658-19"}],"container-title":["Viruses"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4915\/12\/3\/329\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T09:07:41Z","timestamp":1760173661000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4915\/12\/3\/329"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,3,18]]},"references-count":49,"journal-issue":{"issue":"3","published-online":{"date-parts":[[2020,3]]}},"alternative-id":["v12030329"],"URL":"https:\/\/doi.org\/10.3390\/v12030329","relation":{"has-preprint":[{"id-type":"doi","id":"10.1101\/723114","asserted-by":"object"}]},"ISSN":["1999-4915"],"issn-type":[{"value":"1999-4915","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,3,18]]}}}